scholarly journals Reproductive failure in mice lacking inter-alpha-trypsin inhibitor (ITI) – ITI target genes in mouse ovary identified by microarray analysis

2004 ◽  
Vol 183 (1) ◽  
pp. 29-38 ◽  
Author(s):  
Mika Suzuki ◽  
Hiroshi Kobayashi ◽  
Yoshiko Tanaka ◽  
Naohiro Kanayama ◽  
Toshihiko Terao
Keyword(s):  
Real Time ◽  
Trypsin Inhibitor ◽  
Target Genes ◽  
Reproductive Function ◽  
Pcr Analysis ◽  
Rt Pcr ◽  
Reproductive Process ◽  
Female Mice ◽  
Retinoid Metabolism ◽  
Regulatory Effects

Bikunin, a Kunitz-type protease inhibitor, is found in blood and urine. It has been established by two laboratories independently that the bikunin knockout female mice display a severe reduction in fertility: the cumulus oophorus has a defect in forming the extracellular hyaluronan-rich matrix during expansion. Proteins of the inter-alpha-trypsin inhibitor (ITI) family are eliminated in mice in which the bikunin gene has been inactivated, since bikunin is essential for their biosynthesis. Proteins of the ITI family may contribute to the microenvironment in which ovulation takes place. It is not clear, however, whether a single mechanism affects the reproductive function including ovulation. For identifying the full repertoire of the ITI deficiency-related genes, a cDNA microarray hybridization screening was conducted using mRNA from ovaries of wild-type or bik−/− female mice. A number of genes were identified and their regulation was confirmed by real-time RT-PCR analysis. Our screen identified that 29 (0.7%) and 5 genes (0.1%) of the genes assayed were, respectively, up- and down-regulated twofold or more. The identified genes can be classified into distinct subsets. These include stress-related, apoptosis-related, proteases, signaling molecules, aging-related, cytokines, hyaluronan metabolism and signaling, reactive oxygen species-related, and retinoid metabolism, which have previously been implicated in enhancing follicle development and/or ovulation. Real-time RT-PCR analysis confirmed that these genes were up- and down-regulated two- to tenfold by bikunin knockout. These studies demonstrate that proteins of the ITI family may exert potent regulatory effects on a major physiological reproductive process, ovulation.

Real-time RT-PCR analysis of housekeeping genes in human skeletal muscle following acute exercise

2004 ◽  
Vol 18 (2) ◽  
pp. 226-231 ◽  
Author(s):  
Douglas J. Mahoney ◽  
Kate Carey ◽  
Ming-Hua Fu ◽  
Rodney Snow ◽  
David Cameron-Smith ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Real Time ◽  
Healthy Subjects ◽  
Acute Exercise ◽  
Housekeeping Genes ◽  
Housekeeping Gene ◽  
Cycle Ergometry ◽  
Pcr Analysis ◽  
28S Rrna ◽  
Rt Pcr

Studies examining gene expression with RT-PCR typically normalize their mRNA data to a constitutively expressed housekeeping gene. The validity of a particular housekeeping gene must be determined for each experimental intervention. We examined the expression of various housekeeping genes following an acute bout of endurance (END) or resistance (RES) exercise. Twenty-four healthy subjects performed either a interval-type cycle ergometry workout to exhaustion (∼75 min; END) or 300 single-leg eccentric contractions (RES). Muscle biopsies were taken before exercise and 3 h and 48 h following exercise. Real-time RT-PCR was performed on β-actin, cyclophilin (CYC), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and β2-microglobulin (β2M). In a second study, 10 healthy subjects performed 90 min of cycle ergometry at ∼65% of V̇o2 max, and we examined a fifth housekeeping gene, 28S rRNA, and reexamined β2M, from muscle biopsy samples taken immediately postexercise. We showed that CYC increased 48 h following both END and RES exercise (3- and 5-fold, respectively; P < 0.01), and 28S rRNA increased immediately following END exercise (2-fold; P = 0.02). β-Actin trended toward an increase following END exercise (1.85-fold collapsed across time; P = 0.13), and GAPDH trended toward a small yet robust increase at 3 h following RES exercise (1.4-fold; P = 0.067). In contrast, β2M was not altered at any time point postexercise. We conclude that β2M and β-actin are the most stably expressed housekeeping genes in skeletal muscle following RES exercise, whereas β2M and GAPDH are the most stably expressed following END exercise.

scholarly journals Pioglitazone Attenuates Vascular Fibrosis in Spontaneously Hypertensive Rats

PPAR Research ◽  
2012 ◽  
Vol 2012 ◽  
pp. 1-7 ◽  
Author(s):  
Dengfeng Gao ◽  
Ning Ning ◽  
Guanghua Hao ◽  
Xiaolin Niu
Keyword(s):  
Real Time ◽  
Immunohistochemical Staining ◽  
Spontaneously Hypertensive Rats ◽  
Pcr Analysis ◽  
Rt Pcr ◽  
Hypertensive Rats ◽  
Spontaneously Hypertensive ◽  
Ppar Γ ◽  
Collagen Iii ◽  
Wistar Kyoto

Objective. We sought to investigate whether the peroxisome proliferator-activated receptor-γ (PPAR-γ) ligand pioglitazone can attenuate vascular fibrosis in spontaneously hypertensive rats (SHRs) and explore the possible molecular mechanisms.Methods. SHRs (8-week-old males) were randomly divided into 3 groups (n=8each) for treatment: pioglitazone (10 mg/kg/day), hydralazine (25 mg/kg/day), or saline. Normal male Wistar Kyoto (WKY) rats (n=8) served as normal controls. Twelve weeks later, we evaluated the effect of pioglitazone on vascular fibrosis by Masson’s trichrome and immunohistochemical staining of collagen III and real-time RT-PCR analysis of collagen I, III and fibronectin mRNA.Vascular expression of PPAR-γ and connective tissue growth factor (CTGF) and transforming growth factor-β (TGF-β) expression were evaluated by immunohistochemical staining, western blot analysis, and real-time RT-PCR.Results. Pioglitazone and hydralazine treatment significantly decreased systolic blood pressure in SHRs. Masson’s trichrome staining for collagen III and real-time RT-PCR analysis of collagen I, III and fibronectin mRNA indicated that pioglitazone significantly inhibited extracellular matrix production in the aorta. Compared with Wistar Kyoto rats, SHRs showed significantly increased vascular CTGF expression. Pioglitazone treatment significantly increased PPAR-γ expression and inhibited CTGF expression but had no effect on TGF-β expression.Conclusions. The results indicate that pioglitazone attenuated vascular fibrosis in SHRs by inhibiting CTGF expression in a TGF-β-independent mechanism.

scholarly journals Early Detection of Melanoma Progression by Quantitative Real-time RT-PCR Analysis for Multiple Melanoma Markers

2008 ◽  
Vol 57 (1) ◽  
pp. 57-64 ◽  
Author(s):  
Peter Arenberger ◽  
Monika Arenbergerova ◽  
Olga Vohradnikova ◽  
Jaromir Kremen
Keyword(s):  
Early Detection ◽  
Real Time ◽  
Pcr Analysis ◽  
Rt Pcr ◽  
Melanoma Progression

Quantitative real-time RT-PCR analysis of eight novel estrogen-regulated genes in breast cancer

2003 ◽  
Vol 18 (2) ◽  
pp. 123-129 ◽  
Author(s):  
V. Sorbello ◽  
L. Fuso ◽  
C. Sfiligoi ◽  
C. Scafoglio ◽  
R. Ponzone ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Real Time ◽  
Pcr Analysis ◽  
Rt Pcr

scholarly journals The Effect of Laminin-1-Doped Nanoroughened Implant Surfaces: Gene Expression and Morphological Evaluation

2012 ◽  
Vol 2012 ◽  
pp. 1-9 ◽  
Author(s):  
Humberto Osvaldo Schwartz-Filho ◽  
Kostas Bougas ◽  
Paulo G. Coelho ◽  
Ying Xue ◽  
Mariko Hayashi ◽  
...  
Keyword(s):  
Gene Expression ◽  
Real Time ◽  
Rabbit Model ◽  
Interleukin 10 ◽  
Control Surface ◽  
Pcr Analysis ◽  
Tartrate Resistant Acid Phosphatase ◽  
Rt Pcr ◽  
Bony Tissue ◽  
Laminin 1

Aim. This study aimed to observe the morphological and molecular effect of laminin-1 doping to nanostructured implant surfaces in a rabbit model.Materials and Methods. Nanostructured implants were coated with laminin-1 (test; dilution, 100 μg/mL) and inserted into the rabbit tibiae. Noncoated implants were used as controls. After 2 weeks of healing, the implants were removed and subjected to morphological analysis using scanning electron microscopy (SEM) and gene expression analysis using the real-time reverse transcriptase-polymerase chain reaction (RT-PCR).Results. SEM revealed bony tissue attachment for both control and test implants. Real-time RT-PCR analysis showed that the expression of osteoblast markers RUNX-2, osteocalcin, alkaline phosphatase, and collagen I was higher (1.62-fold, 1.53-fold, 1.97-fold, and 1.04-fold, resp.) for the implants modified by laminin-1 relative to the control. All osteoclast markers investigated in the study presented higher expression on the test implants than controls as follows: tartrate-resistant acid phosphatase (1.67-fold), calcitonin receptor (1.35-fold), and ATPase (1.25-fold). The test implants demonstrated higher expression of inflammatory markers interleukin-10 (1.53-fold) and tumour necrosis factor-α(1.61-fold) relative to controls.Conclusion. The protein-doped surface showed higher gene expression of typical genes involved in the osseointegration cascade than the control surface.

Real-time RT-PCR analysis of primary and metastatic colon cancer for the presence of JC virus T-antigen

2006 ◽  
Vol 130 (2) ◽  
pp. 261
Author(s):  
P. Sturrock ◽  
E. Tsai ◽  
J. Pitcher ◽  
R.K. Yantiss ◽  
A. Growney ◽  
...  
Keyword(s):  
Colon Cancer ◽  
Real Time ◽  
Jc Virus ◽  
T Antigen ◽  
Pcr Analysis ◽  
Metastatic Colon Cancer ◽  
Rt Pcr

scholarly journals REAL TIME POLYMERASE CHAIN REACTION (RT-PCR) FOR DNA PIG CONTAMINATION IDENTIFICATION SAUSAGE IN SIX DISTRICT PANDEGLANG BANTEN

2021 ◽  
Vol 1 (1) ◽  
pp. 81-88
Author(s):  
Hadi Susilo
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Meat Product ◽  
Meat Products ◽  
Pcr Analysis ◽  
Processed Meat ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Dna Purity ◽  
Polymerase Chain

Sausage is a meat product processed that is popular food especially in Pandeglang, Banten Province. The importance of halal certificates or the existence of the MUI (Indonesian Ulama Council) halal logo for processed meat products makes Muslim people confident to consume them. The aim this research was to identify pig DNA contamination in sausage products in six  districts in Pandeglang without the MUI halal labels using RT-PCR (Real Time-Polymerase Chain Reaction). RT PCR that can calculate to pig to fill these sample free from pig contamination. This research was divided into two stage, the first stage is extracted or carried out DNA and the second stage is RT PCR analysis. The results of the DNA purity test on sausage samples had DNA purity values ​​of 1.84-1.9 (A260 / A280) and resulted in sample concentrations ranging from 37.8 to 102.5 ng / µl.  The only amplification on the FAM curve was in the positive control pig.  the Cq value ranges from 30 - 31.29. The results of RT PCR on sausage samples in the district area in Pandeglang Banten did not detect the presence of pig DNA.

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